Abstract
Cellular reprogramming of committed cells into a pluripotent state can be induced by ectopic expression of genes such as OCT4, SOX2, KLF4, and MYC. Reprogrammed cells can be maintained by activating endogenous pluripotent networks without transgene expression. Although various research groups have attempted to generate pig induced pluripotent stem cells (iPSCs), authentic iPSCs have not be obtained, instead showing dependence on transgene expression. In this study, iPSCs were derived from porcine fetal fibroblasts via drug-inducible vectors carrying human transcription factors (OCT4, SOX2, KLF4, and MYC). Therefore, this study investigated characteristics of iPSCs and reprogramming mechanisms in pig. The iPSCs were stably maintained over an extended period with potential in vitro differentiation into three germ layers. In addition, the pluripotent state of iPSCs was regulated by modulating culture conditions. They showed naive- or primed-like pluripotent states in LIF or bFGF supplemented culture conditions, respectively. However, iPSCs could not be maintained without ectopic expression of transgenes. The cultured iPSCs expressed endogenous transcription factors such as OCT4 and SOX2, but not NANOG (a known gateway to complete reprogramming). Endogenous genes related to mesenchymal-to-epithelial transition (DPPA2, CDH1, EPCAM, and OCLN) were not sufficiently reactivated, as measured by qPCR. DNA methylation analysis for promoters of OCT4, NANOG, and XIST showed that epigenetic reprogramming did not occur in female iPSCs. Based on our results, expression of exogenous genes could not sufficiently activate the essential endogenous genes and remodel the epigenetic milieu to achieve faithful pluripotency in pig. Accordingly, investigating iPSCs could help us improve and develop reprogramming methods by understanding reprogramming mechanisms in pig.
Highlights
Pluripotent stem cells (PSCs) are a promising tool for human regenerative medicine, and in domestic animals, as useful tools for producing transgenic and disease model animals
Because gene expression is regulated by drugs, drug-inducible vector systems have been used for induced pluripotent stem cells (iPSCs) studies on the generation of iPSCs, secondary iPS systems [31], reprogramming mechanisms [11, 12], and inducing naïve pluripotent stem cells in humans [2, 3]
Pig iPSCs were stably maintained over an extended period, and could differntiate into three germ layers
Summary
Pluripotent stem cells (PSCs) are a promising tool for human regenerative medicine, and in domestic animals, as useful tools for producing transgenic and disease model animals. Compared with the primed state, naïve PSCs have developmental and functional ground states showing contributions to blastocyst chimeras and higher transgenic efficiency [2, 3]. For these reasons, many research groups have attempted to generate naïve PSCs of domestic animals (especially pig) for the efficient production of transgenic animals [4,5,6,7]
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
